Medial-Golgi retention of N-acetylglucosaminyltransferase I. Contribution from all domains of the enzyme.
Medial-Golgi retention of N-acetylglucosaminyltransferase I. Contribution from all domains of the enzyme.
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N-乙酰氨基葡萄糖转移酶 I 的内侧高尔基体保留。来自该酶所有域的贡献。
DOI：
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发表时间：
1994
影响因子：
4.8
通讯作者：
P. Gleeson
中科院分区：
文献类型：
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作者：
J. Burke;J. Pettitt;D. Humphris;P. Gleeson
我们之前已经证明，β-1,2-N-乙酰氨基葡萄糖转移酶 I (GnTI) 的跨膜结构域和侧翼残基可以将杂合分子定位到内侧高尔基体池 (Burke, J.、Pettit, J. M.、Schachter, H.、Sarkar, M. 和 Gleeson, P.A. (1992) J. Biol. Chem. 267， 24433-24440）。在这里，我们通过分析小鼠细胞中稳定表达的杂合分子的定位，进一步研究了 GnTI 的胞质尾部、跨膜结构域和催化活性管腔结构域在内侧高尔基体定位中的贡献。与有效定位于内侧高尔基体且在细胞表面未检测到的野生型 GnTI 相比，含有 GnTI 三个结构域中任意两个的杂合分子定位于内侧高尔基体，并且在细胞表面也以低水平存在。与含有两个 GnTI 结构域的分子相比，仅含有 GnTI 跨膜结构域或管腔结构域的杂合分子显示出部分高尔基体保留以及细胞表面表达水平增加。细胞质尾部独立地无法保留高尔基体的报告序列，但增加了含有 GnTI 的腔或跨膜结构域的构建体定位到高尔基体的能力。因此，GnTI 的所有三个结构域对内侧高尔基体定位都有显着贡献。此外，细胞表面表达增加的 GnTI 杂合分子更容易在低盐缓冲液中提取，这表明高尔基体局部 GnTI 的理化特性与细胞表面 GnTI 不同。基于定位的聚集模型，我们提出这些杂合分子的高尔基体保留是通过它们的 GnTI 结构域与转染细胞高尔基体膜内内源糖基转移酶聚集体的相应结构域的相互作用介导的。
We have previously shown that the transmembrane domain and flanking residues of beta-1,2-N-acetylglucosaminyltransferase I (GnTI) can localize a hybrid molecule to medial-Golgi cisternae (Burke, J., Pettitt, J. M., Schachter, H., Sarkar, M., and Gleeson, P.A. (1992) J. Biol. Chem. 267, 24433-24440). Here, we have further examined the contribution of the cytoplasmic tail, transmembrane domain, and the catalytically active, luminal domain of GnTI in medial-Golgi localization, by analyzing the localization of hybrid molecules stably expressed in murine cells. In contrast to wild-type GnTI, which was efficiently localized to the medial-Golgi and not detected at the cell surface, hybrid molecules containing any two of the three domains of GnTI were localized to the medial-Golgi and were also present at low levels at the cell surface. Hybrid molecules containing only the transmembrane domain or the luminal domain of GnTI showed partial Golgi retention together with an increased level of cell surface expression compared with molecules containing two GnTI domains. The cytoplasmic tail independently was unable to retain reporter sequences to the Golgi but increased the ability of constructs containing either the luminal or transmembrane domain of GnTI to localize to the Golgi apparatus. Therefore, all three domains of GnTI contribute significantly to medial-Golgi localization. Furthermore, GnTI hybrid molecules showing increased cell surface expression were more readily extracted in a low salt buffer, suggesting that Golgi localized GnTI differs in physicochemical properties from cell surface GnTI. Based on an aggregation model of localization, we propose that Golgi retention of these hybrid molecules is mediated by the interaction of their GnTI domains with the corresponding domains of endogenous glycosyltransferase aggregates within the Golgi membranes of the transfected cell.